Project Summary Uterine fibroids (also known as leiomyomas) are the most common tumors in the female reproductive tract. Some estimates indicate that up to 75% of American women have uterine fibroids, depending on race and ethnicity. There is a significant disparity in the incidence of fibroids since African-American women are 2-3 times more likely to develop fibroids, often with earlier onset and greater severity of symptoms. These tumors can be very painful and are the leading cause of hysterectomies in the US. Despite the healthcare burden caused by uterine fibroids, their etiology and pathophysiology are unknown. The vast majority of fibroids either have mutations in Mediator Complex protein 12 (MED12) gene or over-express high mobility group AT hooks (HMGA)1/2 transcription factors, but the molecular mechanism that are disrupted and lead to tumor development or growth are unknown. Another well known characteristic of fibroids is that they are mostly clonal, indicating a single cell of origin for each tumor. Our overarching hypothesis is that the vast majority of uterine fibroids develop from myometrial stem/progenitor cells that have become dysregulated by either MED12 mutation or HMGA1/2 over-expression. Using SUSD2 as a marker, we have enriched for a population of perivascular myometrial cells with all the characteristics of mesenchymal stem cells from both human myometrial and fibroid specimens. The perivascular niche is the most common site for mesenchymal stem cells in many tissues. We have compelling evidence that chromatin compartmentalization is changed in uterine fibroids compared to normal myometrium, which leads to homeotic transformation of the stem cell into a more cervical phenotype. We propose to determine how the epigenetic landscape of the myometrial and fibroid stem cells affects compartmentalization in the stem cells. Once these mechanisms are understood, we will have a better understanding of how disruption of key pathways affect their behavior, with the ultimate goal of identifying targets for therapeutic intervention.